CD9 is a glycoprotein receptor belonging to tetraspanin family having molecular weight about 24-27 kD and is known to regulate signal transduction events playing important roles in development, activity, growth and motility of a cell. In addition, CD9 is known being capable of triggering platelet activation and aggregation which regulate cell attachment (Anton, E. S., et al., J. Neurosci. 15:584-595, 1995) and cell migration (Klein-Soyer, C., et al., Arterioscler Thromb Vasc Biol. 20:360-369, 2000). Moreover, it is known to be involved in various cellular phenomena such as promotion of muscle cell fusion and myotube maintenance.
Tetraspanin family such as CD9 has 4 trans-membrane segments and N- and C-terminal thereof exists in intracellular side CD9. In this alignment model, two extracellular loops (ECLs) protrude between the 1st and the 2nd and between the 3rd and the 4th trans-membrane segments, respectively. Thus, tetraspanins are related to various cellular procedures and their various functions seem to be related to their abilities acting as molecular facilitators. The tetraspanins are known to interact with their partner molecules such as some integrins as well as same tetraspanin family molecules such as CD81 and CD63 and the interaction is so called ‘Tetraspanin web’ (Radford, K. J., et al., Biochem. Biophys. Res. Comm. 222:13-18, 1996; Iwamoto, R., et al.,; Le Naour, F., et al., Mol. Cell. Proteomics 5:845-857, 2006).
There is a report that the 2nd extracellular loop (ECL2) is important to cell attachment (George, A., et al., Blood 100: 4502-4511, 2002). In addition, there is a report that in other tetraspanin family molecules, ECL2 domain is glycosylated, although in CD9 ECL1 domain is glycosylated and ECL2 of CD9 is important to promote activity of Diphteria toxin receptor (DTR) toward Diphteria toxin (DT) (Hidetoshi, H., et al. 289:782-790, 2001). Besides the reports, many researchers are interested in the function of ECL2 of CD9, but the function is not understood clearly yet.
With respect to cancer, CD9 is called as “motility-related antigen, MRP-1” and is reported to be related with cell motility and tumor metastasis (Miyake, M. and Hakomori, S., Biochemistry 30:3328-3334, 1991). However, it is controversial since regarding the role of CD9 in cancer there are some reports showing contrary results according to type of cancers. For example, decreases of expression of CD9 are reported in colon cancer (Mori, M., et al., Clin. Cancer Res. 4:1507-1510, 1998), breast cancer (Miyake, M., et al., Cancer Res. 55:4127-4131, 1995), lung cancer (Higachiyama, M., et al., Cancer Res. 55:6040-6044, 1995; Funakoshi, T., et al., Oncogene 22:674-687, 2003) and pancreatic cancer (Sho, M., et al., Int. J. Cancer 79:509-516, 1998) and it is reported that this is associated with invasion, metastasis and poor prognosis of patients. However, there are some reports that expression of CD9 is increased in head and neck squamous cell carcinoma (Erovic, B. M., et al., Head Neck 25:848-857, 2003) and stomach cancer (Hori, H., et al., J. Surg. Res. 117:208-215, 2004) according to the progression of cancers. These contrary reports draw a deduction that CD9 has tissue-specific aspects. Microarray or immunohistochemistry assays for CD9 in ovarian cancers are reported (Drapkin, R., et al., Hum Pathol. 35:1014-1021, 2004; Peters, D. G., et al., Cancer Epidemiol Biomarkers Prev. 14:1717-1723, 2005; Houle, C. D., et al., Gynecol Oncol 86:69-78, 2002), but no function thereof in ovarian is reported.
Based on the fact that poor prognosis of ovarian cancer patients is related to CD9 over-expression, the present inventors tried to develop a CD9 specific antibody. And as a result, the present inventors completed this invention by confirming that human antibodies 10E4, 11G, 3F3, 8A, 12F and 5G4 recognize CD9 extracellular loop 2 domain (CD9-ECL2) as an epitope and thus strongly bind to CD9, are capable of neutralizing CD9, and inhibiting proliferation, invasion and migration of cancer cells.